Serum starvation and recovery

Serum starvation was performed overnight (18 h) and recovered by resupplementation of media containing 10% FCS for 10 min. Where indicated, amino acid starvation was performed for 1 h (after one wash with PBS) in RPMI 1640 without amino acids (US Biologicals), and recovery was achieved by addition of 1× RPMI 1640 amino acid solution (R7131; Sigma-Aldrich) with or without 10% FCS, pH 7.2, for 10 min. Where indicated, cells were incubated for the last 2 h before harvest/fixation with the ROCKi Y-27632 (50 µM; SCM075; Sigma-Aldrich) or the integrin-binding peptide cilengitide (5 µM; SML1594; Sigma-Aldrich). Where indicated, cells were preincubated with 100 nM rapamycin for 2 h.